Regeneration of starfish radial nerve cord restores animal mobility and unveils a new coelomocyte population.

The potential to regenerate a damaged body part is expressed to a different extent in animals. Echinoderms, in particular starfish, are known for their outstanding regenerating potential. Differently, humans have restricted abilities to restore organ systems being dependent on limited sources of stem cells. In particular, the potential to regenerate the central nervous system is extremely limited, explaining the lack of natural mechanisms that could overcome the development of neurodegenerative diseases and the occurrence of trauma. Therefore, understanding the molecular and cellular mechanisms of regeneration in starfish could help the development of new therapeutic approaches in humans. In this study, we tackle the problem of starfish central nervous system regeneration by examining the external and internal anatomical and behavioral traits, the dynamics of coelomocyte populations, and neuronal tissue architecture after radial nerve cord (RNC) partial ablation. We noticed that the removal of part of RNC generated several anatomic anomalies and induced behavioral modifications (injured arm could not be used anymore to lead the starfish movement). Those alterations seem to be related to defense mechanisms and protection of the wound. In particular, histology showed that tissue patterns during regeneration resemble those described in holothurians and in starfish arm tip regeneration. Flow cytometry coupled with imaging flow cytometry unveiled a new coelomocyte population during the late phase of the regeneration process. Morphotypes of these and previously characterized coelomocyte populations were described based on IFC data. Further studies of this new coelomocyte population might provide insights on their involvement in radial nerve cord regeneration.

Cell Uptake of Steroid-BODIPY Conjugates and Their Internalization Mechanisms: Cancer Theranostic Dyes.

Estradiol-BODIPY linked via an 8-carbon spacer chain and 19-nortestosterone- and testosterone-BODIPY linked via an ethynyl spacer group were evaluated for cell uptake in the breast cancer cell lines MCF-7 and MDA-MB-231 and prostate cancer cell lines PC-3 and LNCaP, as well as in normal dermal fibroblasts, using fluorescence microscopy. The highest level of internalization was observed with 11ß-OMe-estradiol-BODIPY 2 and 7α-Me-19-nortestosterone-BODIPY 4 towards cells expressing their specific receptors. Blocking experiments showed changes in non-specific cell uptake in the cancer and normal cells, which likely reflect differences in the lipophilicity of the conjugates. The internalization of the conjugates was shown to be an energy-dependent process that is likely mediated by clathrin- and caveolae-endocytosis. Studies using 2D co-cultures of cancer cells and normal fibroblasts showed that the conjugates are more selective towards cancer cells. Cell viability assays showed that the conjugates are non-toxic for cancer and/or normal cells. Visible light irradiation of cells incubated with estradiol-BODIPYs 1 and 2 and 7α-Me-19-nortestosterone-BODIPY 4 induced cell death, suggesting their potential for use as PDT agents.

Doxorubicin-sensitive and -resistant colorectal cancer spheroid models: assessing tumor microenvironment features for therapeutic modulation.

Introduction: The research on tumor microenvironment (TME) has recently been gaining attention due to its important role in tumor growth, progression, and response to therapy. Because of this, the development of three-dimensional cancer models that mimic the interactions in the TME and the tumor structure and complexity is of great relevance to cancer research and drug development. Methods: This study aimed to characterize colorectal cancer spheroids overtime and assess how the susceptibility or resistance to doxorubicin (Dox) or the inclusion of fibroblasts in heterotypic spheroids influence and modulate their secretory activity, namely the release of extracellular vesicles (EVs), and the response to Dox-mediated chemotherapy. Different characteristics were assessed over time, namely spheroid growth, viability, presence of hypoxia, expression of hypoxia and inflammation-associated genes and proteins. Due to the importance of EVs in biomarker discovery with impact on early diagnostics, prognostics and response to treatment, proteomic profiling of the EVs released by the different 3D spheroid models was also assessed. Response to treatment was also monitored by assessing Dox internalization and its effects on the different 3D spheroid structures and on the cell viability. Results and Discussion: The results show that distinct features are affected by both Dox resistance and the presence of fibroblasts. Fibroblasts can stabilize spheroid models, through the modulation of their growth, viability, hypoxia and inflammation levels, as well as the expressions of its associated transcripts/proteins, and promotes alterations in the protein profile exhibit by EVs. Summarily, fibroblasts can increase cell-cell and cell-extracellular matrix interactions, making the heterotypic spheroids a great model to study TME and understand TME role in chemotherapies resistance. Dox resistance induction is shown to influence the internalization of Dox, especially in homotypic spheroids, and it is also shown to influence cell viability and consequently the chemoresistance of those spheroids when exposed to Dox. Taken together these results highlight the importance of finding and characterizing different 3D models resembling more closely the in vivo interactions of tumors with their microenvironment as well as modulating drug resistance.

Combined cancer therapeutics-Tackling the complexity of the tumor microenvironment.

Cancer treatment has yet to find a "silver bullet" capable of selectively and effectively kill tumor cells without damaging healthy cells. Nanomedicine is a promising field that can combine several moieties in one system to produce a multifaceted nanoplatform. The tumor microenvironment (TME) is considered responsible for the ineffectiveness of cancer therapeutics and the difficulty in the translation from the bench to bed side of novel nanomedicines. A promising approach is the use of combinatorial therapies targeting the TME with the use of stimuli-responsive nanomaterials which would increase tumor targeting. Contemporary combined strategies for TME-targeting nanoformulations are based on the application of external stimuli therapies, such as photothermy, hyperthermia or ultrasounds, in combination with stimuli-responsive nanoparticles containing a core, usually composed by metal oxides or graphene, and a biocompatible stimuli-responsive coating layer that could also contain tumor targeting moieties and a chemotherapeutic agent to enhance the therapeutic efficacy. The obstacles that nanotherapeutics must overcome in the TME to accomplish an effective therapeutic cargo delivery and the proposed strategies for improved nanotherapeutics will be reviewed. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease Nanotechnology Approaches to Biology > Nanoscale Systems in Biology Therapeutic Approaches and Drug Discovery > Emerging Technologies.

Cu(I) complexes as new antiproliferative agents against sensitive and doxorubicin resistant colorectal cancer cells: synthesis, characterization, and mechanisms of action.

Cancer is one of the worst health issues worldwide, representing the second leading cause of death. Current chemotherapeutic drugs face some challenges like the acquired resistance of the tumoral cells and low specificity leading to unwanted side effects. There is an urgent need to develop new compounds that may target resistant cells. The synthesis and characterization of two Cu(i) complexes of general formula [Cu(PP)(LL)][BF4], where PP is a phosphane ligand (triphenylphosphine or 1,2-bis(diphenylphosphano) ethane) and LL = is a heteroaromatic bidentate ligand (4,4'-dimethyl-2,2'-bipyridine and 6,3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine). The new compounds were fully characterized by spectroscopic techniques (NMR, FTIR and UV-vis.), elemental analysis (C, H, N and S) and two structures were determined by single X-ray diffraction studies. The antiproliferative potential of the new Cu(i) complexes were studied in tumor (breast adenocarcinoma, ovarian carcinoma and in colorectal carcinoma sensitive and resistant to doxorubicin) and normal (fibroblasts) cell lines. Complexes 1-4 did not show any antiproliferative potential. Amongst the complexes 5-8, complex 8 shows high cytotoxic potential against colorectal cancer sensitive and resistant to doxorubicin and low cytotoxicity towards healthy cells. We show that complexes 5-8 can cleave pDNA and, in particular, the in vitro pDNA cleavage is due to an oxidative mechanism. This oxidative mechanism corroborates the induction of reactive oxygen species (ROS), that triggers HCT116 cell death via apoptosis, as proved by the increased expression of BAX protein relative to BCL-2 protein and the depolarization of mitochondrial membrane potential, and via autophagy. Additionally, complex 8 can block the cell cycle in the G1 phase, also exhibiting a cytostatic potential. Proteomic analysis confirmed the apoptotic, autophagic and cytostatic potential of complex 8, as well as its ability to produce ROS and cause DNA damage. The interference of the complex in folding and protein synthesis and its ability to cause post-translational modifications was also verified. Finally, it was observed that the complex causes a reduction in cellular metabolism. The results herein demonstrated the potential of Cu(i) complexes in targeting doxorubicin sensitive and resistant cells which is positive and must be further explored using in vivo animal models.